Optimizing mesoderm progenitor selection and three-dimensional microniche culture allows highly efficient endothelial differentiation and ischemic tissue repair from human pluripotent stem cells

Abstract

BackgroundGeneration of large quantities of endothelial cells is highly desirable for vascular research, for the treatment of ischemia diseases, and for tissue regeneration. To achieve this goal, we developed a simple, chemically defined culture system to efficiently and rapidly differentiate endothelial cells from human pluripotent stem cells by going through an MESP1 mesoderm progenitor stage.MethodsMesp1 is a key transcription factor that regulates the development of early cardiovascular tissue. Using an MESP1-mTomato knock-in reporter human embryonic stem cell line, we compared the gene expression profiles of MESP1+ and MESP1− cells and identified new signaling pathways that may promote endothelial differentiation. We also used a 3D scaffold to mimic the in vivo microenvironment to further improve the efficiency of endothelial cell generation. Finally, we performed cell transplantation into a critical limb ischemia mouse model to test the repairing potential of endothelial-primed MESP1+ cells.ResultsMESP1+ mesoderm progenitors, but not MESP1− cells, have strong endothelial differentiation potential. Global gene expression analysis revealed that transcription factors essential for early endothelial differentiation were enriched in MESP1+ cells. Interestingly, MESP1 cells highly expressed Sphingosine-1-phosphate (S1P) receptor and the addition of S1P significantly increased the endothelial differentiation efficiency. Upon seeding in a novel 3D microniche and priming with VEGF and bFGF, MESP1+ cells markedly upregulated genes related to vessel development and regeneration. 3D microniches also enabled long-term endothelial differentiation and proliferation from MESP1+ cells with minimal medium supplements. Finally, we showed that transplanting a small number of endothelial-primed MESP1+ cells in 3D microniches was sufficient to mediate rapid repair of a mouse model of critical limb ischemia.ConclusionsOur study demonstrates that combining MESP1+ mesoderm progenitor cells with tissue-engineered 3D microniche and a chemically defined endothelial induction medium is a promising route to maximizing the production of endothelial cells in vitro and augment their regenerative power in vivo.Electronic supplementary materialThe online version of this article (doi:10.1186/s13287-016-0455-4) contains supplementary material, which is available to authorized users.

Transferrin improved the generation of cardiomyocyte from human pluripotent stem cells for myocardial infarction repair

“…H1, H9 hESCs (WiCell Institute), HUES8 hESCs (Harvard Stem Cell Institute), and human induced pluripotent stem cells (CD34-iPSCs Zhang et al 2017 ; Duan et al 2018 ), provided by Dr. J. Na, Tsinghua University) were routinely maintained on Matrigel (BD Biosciences)—coated plates (Corning) in TeSR-E8 medium (STEMCELL Technologies). hPSCs were passaged every 4 days using Accutase (Merck) and 10 μM ROCK inhibitor Y27632 (Selleck) was added for the first 24 h after passaging.…”

Dgcr8 deletion in the primitive heart uncovered novel microRNA regulating the balance of cardiac-vascular gene program

“…S1C). Nfatc1 was specifically expressed in the endocardium of early developing heart at E9.5, and its expression level decreases at later embryonic and neonatal stages (Zhang et al, 2017 ). We found that Nfatc1 is expressed in the inner layer of E9.5 cKO embryonic hearts, and Pecam1 could also be obviously detected in the E9.5 cKO heart, which means both endocardium and ECs were also exist in the cKO heart (Fig.…”

“…The effects of miRNAs on mouse or human EC function were tested by vascular tube formation and scratch-wound healing assays as described in published studies (He et al, 2008 ; Zhang et al, 2017 ). In brief, for vascular tube formation assay of mouse SVEC4–10 cells (ATCC CRL-2181 TM ), 24-well plate were pre-coated with 150 μL/well Matrigel (1:1, BD Biosciences).…”

Integrative Analysis of DNA Methylation and Gene Expression Profiles Identifies Colorectal Cancer-Related Diagnostic Biomarkers

“…DNA methylation and gene expression profiles of the TCGA-COAD dataset which contained 407 CRC and 46 adjacent samples were downloaded from TCGA and used as a training set. The testing set consisted of two independent DNA methylation datasets, including GSE79740 [ 12 ] which contained 44 CRC samples and 10 normal samples, and GSE42752 [ 13 ] which contained 22 CRC and 41 normal samples from GEO.…”